The optical spectra of chromophores in proteins are broad due to large inhomogeneous broadening under the usual conditions of measurement. Using high resolution laser spectroscopy at cryogenic temperatures to photoselect among conformational substates, resolved spectra are obtained. We have shown that in metal free and metal substituted heme proteins the fluorescence line narrowing and hole burning spectra reveal details of the porphyrin electronic and vibrational structural features and which can be used to compare the vibrational structural features of both the ground and excited state molecules. In the proposed work we will study porphyrin interactions with the polypeptide chain using cytochrome c and myoglobin. The specific aims are to: l. Examine parameters of the heme pocket that determine the porphyrin spectral properties. Cytochrome c and myoglobin from various sources will be selected where specific amino acid changes in the heme pocket allow us to ask whether particular amino acids affect the spectral characteristics. Distribution in the electronic energies will be related to the distribution of conformational substates. 2. Investigate how changes in the spectra can be used to detect changes in the peptide chain, such as those induced by complex formation (as for cyt c binding to CCP) or by altering medium conditions. 3. Examine how the protein polypeptide chain is affected by photoreactions of the porphyrin and consider how the chain influences rates of a photo- tautomerization reaction. As a monitor occurring in the heme pocket we will use high resolution absorption techniques of fluorescence line-narrowing, hole burning and infrared spectroscopies. The aims will be addressed by systematically comparing different metal derivatives, different porphyrins and different proteins and by altering experimental conditions.